Endometriosis is an estrogen-dependent and progesterone-resistant inflammatory disease of reproductive-age women characterized by the presence of functional endometrium outside the uterine cavity. The major symptoms of this enigmatic disease are painful menstruation, chronic pelvic pain, and infertility. Thus, it represents the single major cause for hysterectomy in reproductive-age women in United States, with annual estimated societal cost of ~$69.4 billion. Current anti-estrogen therapies can be prescribed only for a short time because of undesirable side effects and failure to prevent recurrence. Identification of novel signaling pathways that inhibit growth and survival of endometriotic lesions is needed to develop new molecular-targeted therapies for endometriosis. Concentrations of prostaglandin E2 (PGE2) in the peritoneal fluid are higher in endometriosis women compared to disease-free women, and this increased PGE2 is thought to promote the survival, growth, and inflammation of endometriosis. Expression of miR15a and miR34c appears to be epigenetically silenced in endometriotic lesions in human patients. Our overarching hypothesis is that selective inhibition of PGE2 receptors EP2 and EP4 reverses epigenetic silencing of miR15a and miR34c by altering DNA methylation and histone modifications in endometriosis and that combinatorial inhibition of EP2 and EP4 and activation of miR15a and miR34c cooperatively and synergistically inhibit growth and survival of endometriosis. Specific Aim-1 will determine the combinatorial therapeutic effects of selective inhibition of PGE2 receptors EP2 and EP4 and activation of miR15a and miR34c on growth and survival of endometriosis. Specific Aim-2 will identify the molecular and cellular mechanisms through which miR15a and miR34c mediate inhibitory effects of PGE2 receptors EP2 and EP4 on growth and survival of endometriosis. Specific Aim-3 will establish the molecular mechanisms and epigenetic pathways through which inhibition of PGE2 receptors EP2 and EP4 reverses epigenetic silencing and restores transcriptional activation of miR15a and miR34c in endometriosis. The experimental approaches such as the humanized Rag2?(c) mice models, active human endometriotic epithelial and stromal cells, inhibition of EP2 and EP4 and activation of miR15a and miR34c by pharmacologic and genomic approaches, whole animal bioimaging, and epigenetics, molecular biology and microscopy-based techniques will be used. Successful completion of the research project is expected to provide new fundamental knowledge to formulate personalized phenotype-based combinatorial nonsteroidal therapy through inhibition of EP2 and EP4 and activation of miR15a and miR34c for endometriosis. This novel non-hormonal therapy could provide a more effective alternative treatment to existing hormonal therapies and hold promise for future innovative shifts in endometriosis treatment. Equally important, potential treatments identified in this project can be extrapolated to other inflammatory diseases. This application addresses missions of NICHD on women's reproductive health.